This proposal is based on our recent finding that there is a factor(s) in ascites fluid of rats bearing mammary ascites tumors which is a potent stimulator of prolactin release from pituitary cells in vitro. The objectives of the research are 1) a study of the interactions of the ascites fluid factor(s) (AF) with known regulators of prolactin secretion (TRH, estradiol, dopamine) and 2) preliminary characterization of AF factor(s) responsible for the stimulatory activity. To accomplish these objectives, pituitary cells prepared from rats in different endocrine states will be incubated with AF plus or minus TRH, estradiol or dopamine in serum free medium for 2 hours to 3 days to determine their effectiveness in promoting prolactin secretion. The dynamics of the AF stimulatory response will be assessed using a pituitary cell column perifusion system. Human pituitary cells prepared from either surgical specimens (patients undergoing hypophysectomy for breast cancer, pituitary tumor, etc.) or post mortem tissue will also be challenged with prolactin secretagogues (e.g., TRH, estrogen, AF) in the in vitro assay system. Preliminary characterization of the AF factor(s) will be done by routine biochemical procedures (gel filtration, electrophoresis, etc.). The results from these human and rat studies should prove useful not only in advancing understanding of regulatory mechanisms involved in the control of prolactin cell function in breast cancer, but may have important implications in terms of documenting the existence of feedback interaction from the mammary gland to the anterior pituitary.